The central role that the cystic fibrosis transmembrane conductance regulator (CFTR) plays in gastrointestinal anion transport physiology is evident from the intestinal and pancreatic manifestations of the genetic disease cystic fibrosis (CF). CFFR is essential to normal Cl- and HCO3- secretion across epithelial surfaces. Loss of this function in CF has been closely associated with disease processes such as distal obstructive syndrome, meconium ileus, pancreatic insufficiency, gallbladder disease and duodenal ulceration. Furthermore, the inability of the senescent epithelial cells to unload base may interfere with apoptosis, leading to necrotic cell death and accompanying inflammatory reactions that underlie CF pathogenesis. The present proposal focuses on the role that CFTR plays in mechanisms of Cl- and HCO3- secretion in the duodenum. Although anion secretion across this intestinal segment maintains a critical barrier to gastric effluent, little is known of the electrochemical gradients for anion secretion, the identity of anion exchange proteins that coordinate activity with CFTR, or the distribution of these processes along the crypt-villus axis. Studies will be performed on intact duodenum from genetically-altered CF mouse models because they accurately reproduce human CF intestinal disease. Knockout mouse models of other anion transport proteins will be used to dissect the mechanism of Cl- and HCO3- secretion in a physiological setting. We will test several hypotheses predicted from our proposed model of duodenal anion secretion. Microelectrodes and fluorescent dye markers will be used to investigate the hypothesis that CFTR functions as both a Cl- and HCO3- conductance. RT-PCR and immunoblots of normal and CF duodenal epithelia will be used to identify the effect of CFTR activity on anion transport protein expression along the crypt-villus axis. In addition, membrane vesicle and transepithelial flux studies will be used to test the hypothesis that the anion exchanger AE2 provides an alternative Cl- uptake mechanism for transepithelial Cl- secretion whereas the anion exchanger DRA, possibly driven by carbonic anhydrase, operates in parallel with CFTR for transepithelial HCO3- secretion. The successful completion of our proposed studies will firmly establish the mechanisms involved in Cl- and HCO3- secretion across the duodenum. The results will further define the role of CFTR in these processes and therefore have important implications for our understanding of intestinal pathophysiology in CF disease.